Role of salt in reconstituting photocycle behavior in triton-damaged purple membranes by addition of native lipids

We demonstrated recently that damage to the normal bacteriorhodopsin (BR) photocycle caused by brief exposure to dilute Triton X-LOO, which removes lipids from the membrane, can be repaired by adding back an extract of purple membrane (PM) lipids (Dracheva; et al. FEES Lett. 1996, 386, 209). It was shown that reconstitution with lipids required the presence of a high concentration of NaCl (similar to 0.5 M for half-maximal reconstitution). This paper shows that reconstitution can be achieved with divalent and trivalent cations at much lower concentrations than were required for NaCl (i.e. similar to 7.5 mM for Ca2+ and similar to 0.15 mM for La3+ for half-maximal reconstitution), indicating that negative surface charges must be neutralized to allow the added lipids to reach and be incorporated at the proper locus. We also report here that reconstitution in the absence of added salt can be accomplished by protons alone with an apparent pK near 5. The role of salt involves both charge-screening and specific binding. It was found that, contrary to the prediction of the Gouy-Chapmann equation, the pK(app) for reconstitution rises with increasing salt concentration.